Rittenhouse J, Marcus F
Anal Biochem. 1984 May 1;138(2):442-8. doi: 10.1016/0003-2697(84)90836-4.
Protein samples prepared for sodium dodecyl sulfate-polyacrylamide gel electrophoresis are preferentially cleaved at aspartyl-prolyl peptide bonds upon heating at 110 degrees C. The presence of aspartyl-prolyl peptide bonds in a protein can therefore be detected by gel electrophoresis of heated samples and the resulting peptides mapped. The method of heat cleavage also works well with proteins in bands cut from electrophoresed gels using modified stacking conditions in the second electrophoresis. An immunoblotting procedure for peptide mapping of nanogram quantities of specific proteins in complex mixtures is demonstrated. Peptide maps produced by aspartyl-prolyl peptide bond cleavage of fructose-1,6-bisphosphatases from different sources show the effectiveness of the above techniques and suggest a conservation of aspartyl-prolyl peptide bonds in pig kidney and mouse and rat liver fructose-1,6-bisphosphatases.
为十二烷基硫酸钠-聚丙烯酰胺凝胶电泳制备的蛋白质样品在110℃加热时会优先在天冬氨酰-脯氨酰肽键处裂解。因此，通过对加热样品进行凝胶电泳并对产生的肽段进行图谱分析，可检测蛋白质中天冬氨酰-脯氨酰肽键的存在。热裂解方法对于使用第二次电泳中的改良堆积条件从电泳凝胶切下的条带中的蛋白质也同样适用。本文展示了一种用于复杂混合物中纳克级特定蛋白质肽段图谱分析的免疫印迹程序。来自不同来源的果糖-1,6-二磷酸酶经天冬氨酰-脯氨酰肽键裂解产生的肽段图谱表明了上述技术的有效性，并提示猪肾、小鼠和大鼠肝脏果糖-1,6-二磷酸酶中天冬氨酰-脯氨酰肽键具有保守性。
